Basal-like breast cancer (BLBC) presents with an aggressive clinical history, development of recurrence, distant metastasis, shorter survival, and usually occurs in young women. BLBC has an activated epithelial-mesenchymal transition (EMT) program, which provides cells with the increased phenotypic and cellular plasticity required during embryonic development, tissue remodeling, wound healing and metastasis. Our long-term goal is to discover the molecular circuitry that governs the functional plasticity of BLBC cells in metastasis and to identify molecules that may serve as druggable targets for treating this deadly disease. In the last nine years, we have systematically studied the function of Snail and the molecular mechanism by which Snail represses E-cadherin expression in BLBC. Our studies clearly indicate that Snail levels are regulated predominantly through protein ubiquitination. High levels of Snail, due to protein stabilization, contribute significantly to the increased invasion, therapeutic resistance, CSC-like properties, and tumor recurrence. However, the mechanism by which Snail escapes ubiquitination and degradation remains unknown; uncovering this mechanism is crucially important for the development of novel approaches to treat BLBC. Recently, we discovered that Dub3 is a bona fide deubiquitinase of Snail and contributes to Snail stabilization in breast cancer. Dub3 is evolutionarily conserved from Drosophila to humans. Dub3-knockout downregulated Snail and disrupted gastrulation (an event that requires EMT) in Drosophila. Elevated expression of Dub3 correlates unequivocally with Snail expression in BLBC samples and cell lines. Overexpression of Dub3 in luminal breast tumor MCF7 and T47D cell lines, which contain no detectable endogenous Dub3 and Snail, induces transformation to a basal-like phenotype; whereas, Dub3-knockdown downregulates Snail and suppresses cell migration. Intriguingly, Dub3 was an early response gene that appeared after exposure to inflammatory cytokines. We hypothesize that Dub3 is the long-sought missing molecule that regulates cellular plasticity by controlling the level of Snail during EMT and metastasis. The objective of this proposal is to characterize the function and regulation of the Dub3-Snail axis and explore the clinical value of Dub3 as a prognostic biomarker and a druggable target for treating metastatic breast cancer. Guided by strong preliminary data, we will test this hypothesis by pursuing three specific aims: (1) to delineate the molecular mechanism by which Dub3 controls EMT; (2) to determine the function and regulation of Dub3 in breast cancer; and (3) to define the function and therapeutic efficacy of Dub3 in vivo. Our proposal is innovative and significant, because it will not only uncover a new paradigm that significantly increases our understanding of the regulation of cellular plasticity during tumor progression and metastasis but also leads to the development of specific Dub3 inhibitors for treating metastatic breast cancer.